There has been a growing interest in the manufacture and use of microfluidic systems for the acquisition of chemical and biological information. In particular, when conducted in microfluidic volumes, complicated biochemical reactions may be carried out using very small volumes of liquid. Among other benefits, microfluidic systems increase the response time of reactions, minimize sample volume, and lower reagent consumption. When volatile or hazardous materials are used or generated, performing reactions in microfluidic volumes also enhances safety and reduces disposal quantities.
Traditionally, microfluidic devices have been constructed in a planar fashion using techniques that are borrowed from the silicon fabrication industry. Representative systems are described, for example, in some early work by Manz et al. (Trends in Anal. Chem. (1990) 10(5): 144-149; Advances in Chromatography (1993) 33: 1-66). In these publications, microfluidic devices are constructed by using photolithography to define channels on silicon or glass substrates and etching techniques to remove material from the substrate to form the channels. A cover plate is bonded to the top of the device to provide closure. Miniature pumps and valves can also be constructed to be integral (e.g., within) such devices. Alternatively, separate or off-line pumping mechanisms are contemplated.
More recently, a number of methods have been developed that allow microfluidic devices to be constructed from plastic, silicone or other polymeric materials. In one such method, a negative mold is first constructed, and plastic or silicone is then poured into or over the mold. The mold can be constructed using a silicon wafer (see, e.g., Duffy et al., Analytical Chemistry (1998) 70: 4974-4984; McCormick et. al., Analytical Chemistry (1997) 69: 2626-2630), or by building a traditional injection molding cavity for plastic devices. Some molding facilities have developed techniques to construct extremely small molds. Components constructed using a LIGA technique have been developed at the Karolsruhe Nuclear Research center in Germany (see, e.g., Schomburg et al., Journal of Micromechanical Microengineering (1994) 4: 186-191), and commercialized by MicroParts (Dortmund, Germany). Jenoptik (Jena, Germany) also uses LIGA and a hot-embossing technique. Imprinting methods in PMMA have also been demonstrated (see, Martynova et al., Analytical Chemistry (1997) 69: 4783-4789). However, these techniques do not lend themselves to rapid prototyping and manufacturing flexibility. Additionally, the foregoing references teach only the preparation of planar microfluidic structures. Moreover, the tool-up costs for both of these techniques are quite high and can be cost-prohibitive.
When working with fluids in conventional macrofluidic volumes, tasks such as metering discrete fluid volumes and then combining those volumes is relatively straightforward. In microfluidic volumes, however, such tasks are considerably more difficult. Most, if not all, microfluidic systems require some interface to the conventional macrofluidic world. Using conventional techniques, the smallest volume of fluid that can be generated is a droplet, typically ranging in volume between approximately 1-100 microliters. At the low end of this volumetric range it is extremely difficult to consistently create droplets having a reasonably low volumetric standard deviation. Applications in which fluidic metering accuracy is important include combinatorial chemistry and microfluidic synthesis. In both applications, it would be desirable to combine repeatably accurate discrete fluid volumes. For example, it would be desirable to provide stoichiometric microfluidic volumes of reagents and solvents when performing synthesis, and it would be desirable to provide accurate amounts of sample and diluent when performing serial dilution.
A known method for combining fluids is to dispense fluid droplets from pipet tips into a microtiter plate. However, the utility of such a method is limited for several reasons. To begin with, because a conventional microtiter plate is open to atmosphere, evaporation of fluid following dispensation is an inherent problem, and such dispensing must take place in an ultra-clean environment to avoid undesirable contamination. Further, surfactants are often used in conjunction with pipet tips to increase the accuracy of dispensing small volumes. These surfactants can detrimentally compromise the purity of the fluids to be metered, and it may be very challenging to remove the surfactants and purify the fluids of interest for further use. Additionally, after the fluids of interest are combined in a well plate, if further processing is desired, it can be cumbersome to extract and transfer the fluids elsewhere.
Accordingly, there exists a need for improved systems and methods for combining discrete microscale fluid volumes.